Post-translational modifications in DNA topoisomerase 2α highlight the role of a eukaryote-specific residue in the ATPase domain

Bedez, Claire; Lotz, Christophe; Batisse, Claire; Broeck, Arnaud Vanden; Stote, Roland H.; Howard, Eduardo; Pradeau‐Aubreton, Karine; Ruff, Marc; Lamour, Valérie

doi:10.1038/s41598-018-27606-8

Cited by 30 publications

(36 citation statements)

References 62 publications

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“…1a), 20 mg of the full length hTop2α were obtained from 6L of BHK21 C13-2P cell cultures. About 10 to 15% of the protein sample shows some degradation depending on the protein batch, as previously observed 43 . Western blot analysis using TOP2A antibody PA5-46814 (Thermo Fischer Scientific) shows that the C-terminal domain tends to be cleaved off during protein purification despite the use of protease inhibitors (Supplementary Fig.…”

Section: Methodssupporting

confidence: 81%

“…Interestingly, Serine 1213, located at the end of the linker, has been found to be subjected to mitotic phosphorylation and contributes to localization of the Top2α to the centromere 43–45 . Such post-translational modification could regulate the binding of this CTD portion to the G-segment in order to modulate the relaxation activity of the hTop2α activities during the cell cycle.…”

Section: Discussionmentioning

confidence: 99%

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Structural basis for the allosteric regulation of Human Topoisomerase 2α

Broeck

Lotz

Drillien

et al. 2020

Preprint

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The human type IIA topoisomerases (Top2) are essential enzymes that regulate DNA topology and chromosome organization. The Top2α isoform is a prime target for antineoplastic compounds used in cancer therapy that form ternary cleavage complexes with the DNA. Despite extensive studies, structural information on this large dimeric assembly is limited to the catalytic domains, hindering the exploration of allosteric mechanism governing the enzyme activities and the contribution of its non-conserved C-terminal domain (CTD). Herein we present cryo-EM structures of the entire human Top2α nucleoprotein complex in different conformations solved at subnanometer resolutions. Our data unveils the molecular determinants that fine tune the allosteric connections between the ATPase domain and the DNA binding/cleavage domain. Strikingly, the reconstruction of the DNA-binding/cleavage domain uncovers a linker leading to the CTD, which plays a critical role in modulating the enzyme’s activities and opens perspective for the analysis of post-translational modifications.

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Section: Methodssupporting

confidence: 81%

Section: Discussionmentioning

confidence: 99%

Structural basis for the allosteric regulation of Human Topoisomerase 2α

Broeck

Lotz

Drillien

et al. 2020

Preprint

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“…Many of the key differences between the two isoforms are determined by their distinct CTDs [ 11 , 12 ]. The start of the CTD in human TOP2A has been placed variously at residue 1173 up to amino acid 1244, depending on whether sequence alignment or partial proteolysis, has been used to define it [ 11 , 12 , 13 ]. This region of the protein is required for its nuclear localisation [ 14 , 15 ], has been shown to modulate the enzyme’s activity [ 12 , 16 , 17 ] and to be responsible for the preference of the alpha isoform for relaxing positively-supercoiled DNA [ 18 , 19 , 20 ].…”

Section: Introductionmentioning

confidence: 99%

“…The CTD is rich in residues subject to modification, with multiple sites of phosphorylation, SUMOylation, ubiquitination and acetylation identified [ 13 , 33 ] (PhosphositePlus database url: ). This may partly reflect the disordered and accessible nature of this protein domain [ 34 , 35 , 36 ].…”

Section: Introductionmentioning

confidence: 99%

The Impact of the C-Terminal Region on the Interaction of Topoisomerase II Alpha with Mitotic Chromatin

Antoniou-Kourounioti

Mimmack

Porter

et al. 2019

IJMS

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Type II topoisomerase enzymes are essential for resolving DNA topology problems arising through various aspects of DNA metabolism. In vertebrates two isoforms are present, one of which (TOP2A) accumulates on chromatin during mitosis. Moreover, TOP2A targets the mitotic centromere during prophase, persisting there until anaphase onset. It is the catalytically-dispensable C-terminal domain of TOP2 that is crucial in determining this isoform-specific behaviour. In this study we show that, in addition to the recently identified chromatin tether domain, several other features of the alpha-C-Terminal Domain (CTD). influence the mitotic localisation of TOP2A. Lysine 1240 is a major SUMOylation target in cycling human cells and the efficiency of this modification appears to be influenced by T1244 and S1247 phosphorylation. Replacement of K1240 by arginine results in fewer cells displaying centromeric TOP2A accumulation during prometaphase-metaphase. The same phenotype is displayed by cells expressing TOP2A in which either of the mitotic phosphorylation sites S1213 or S1247 has been substituted by alanine. Conversely, constitutive modification of TOP2A by fusion to SUMO2 exerts the opposite effect. FRAP analysis of protein mobility indicates that post-translational modification of TOP2A can influence the enzyme’s residence time on mitotic chromatin, as well as its subcellular localisation.

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“…Distribution of the post-translational modifications on the structures of Top2a catalytic domains. A: modifications found in normal homeostasis cells (Bedez et al [16] ,. 2018) reported on the ATPase domain structure (PDBID: 1ZXM) (top) and DNA binding/ cleavage domain (PDBID: 5GWK) (bottom).…”

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confidence: 99%

The interplay between DNA topoisomerase 2α post-translational modifications and drug resistance

Lotz¹,

Lamour²

2020

CDR

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The type 2 DNA topoisomerases (Top2) are conserved enzymes and biomarkers for cell proliferation. The catalytic activities of the human isoform Top2a are essential for the regulation of DNA topology during DNA replication, transcription, and chromosome segregation. Top2a is a prominent target for anti-cancer drugs and is highly regulated by post-translational modifications (PTM). Despite an increasing number of proteomic studies, the extent of PTM in cancer cells and its importance in drug response remains largely uncharacterized. In this review, we highlight the different modifications affecting the human Top2a in healthy and cancer cells, taking advantage of the structure-function information accumulated in the past decades. We also overview the regulation of Top2a by PTM, the level of PTM in cancer cells, and the resistance to therapeutic compounds targeting the Top2 enzyme. Altogether, this review underlines the importance of future studies addressing more systematically the interplay between PTM and Top2 drug resistance.

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Post-translational modifications in DNA topoisomerase 2α highlight the role of a eukaryote-specific residue in the ATPase domain

Cited by 30 publications

References 62 publications

Structural basis for the allosteric regulation of Human Topoisomerase 2α

Structural basis for the allosteric regulation of Human Topoisomerase 2α

The Impact of the C-Terminal Region on the Interaction of Topoisomerase II Alpha with Mitotic Chromatin

The interplay between DNA topoisomerase 2α post-translational modifications and drug resistance

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